CN101346383A - Triazole derivatives - Google Patents

Abstract

Novel triazole derivatives are inhibitors of TGF-beta receptor I kinase, for the treatment of tumours.

Description

Triazole derivative

Background of invention

The objective of the invention is to seek new compound, particularly can be used for preparing the new compound of medicine with high value character.

The present invention relates to compound and application of compound, in described application, suppress, regulate and control and/or regulate kinase whose signal transduction, particularly the kinase whose signal transduction of TGF-beta receptor works, and the invention still further relates to the pharmaceutical composition that comprises these compounds and described compound and is used for the treatment of purposes by kinases inductive disease.

Transforming growth factor-beta is the prototype of TGF-beta superfamily, and this family is the multi-functional somatomedin of gang's high conservative, all has critical function in fetal development and adult organism body.In Mammals, differentiated three kinds of TGF-β hypotypes (TGF-β 1,2 and 3), wherein TGF-β 1 is modal hypotype (Kingsley (1994) Genes Dev 8:133-146).TGF-β 3 is only for example expressing in the mesenchymal cell, and TGF-β 1 expresses in mesenchymal cell and epithelial cell.TGF-β is former synthetic as precursor protein, is released into (Derynck (1985) Nature 316:701-705 in the extracellular matrix with inactive form; Bottinger (1996) PNAS 93:5877-5882).Except be cut off be also referred to as latent related peptides (LAP) and with ripe zone keeps related precursor zone, one of conjugated protein four kinds of hypotypes of latent TGF-β (LTBP 1-4) also can combine (Gentry (1988) MolCell Biol 8:4162-4168 with TGF-β, Munger (1997), Kindey Int 51:1376-1382).The activation of nonactive complex body is the necessary step of TGF-β performance biological action, but its mechanism also fails to clarify fully.But for example carry out proteolysis processing yes necessary step (Munger (1997) Kindey Int 51:1376-1382) by plasmin, blood plasma trans-glutaminases or thrombostondin.Activatory part TGF-β mediates its biological action by the I type that is wide expression of three TGF-beta receptors on the film and II receptor and III receptor beta glycan and Endoglin, wherein Endoglin only expresses (Gougos (1990) J Biol Chem 264:8361-8364, Loeps-Casillas (1994) J Cell Biol 124:557-568) in endotheliocyte.

Two kinds of III type TGF-beta receptors all lack the intracellular kinase structural domain that can promote that signal transmits in cell.Because III type TGF-beta receptor all combines with high-affinity with whole three kinds of TGF-β hypotypes, and II type TGF-beta receptor pair is also very high with III receptor bonded ligand affinity, therefore thinks that its biological function is to regulate and control to be used for the utilizability of I type and II type TGF-beta receptor part (Lastres (1996) J Cell Biol 133:1109-1121; Lopes-Casillas (1993) Cell 73:1435-1344).Closely-related I type of structure and II receptor have the serine/threonine kinase structural domain, are responsible for transmitting signal in the endochylema zone.II type TGF-beta receptor combines with TGF-β, and I type TGF-beta receptor adds in this signal transmission complex body then.The serine/threonine structural domain of II receptor has activity on forming, seryl residue that can this complex body of phosphorylation in the GS of so-called I receptor structural domain.This phosphorylation activates the kinases of I receptor, signal mediated factor SMAD albumen in the own phosphorylation cell of present I receptor energy, thus cause intracellular signal transmission (being summarized in Derynck (1997) Biochim Biophys Acta 1333:F105-F150).

The protein of SMAD family is the substrate of whole TGF-'beta ' family receptor kinases.Up to now, differentiated 8 SMAD albumen, it can be divided into 3 groups: (1) acceptor relationship type SMAD (R-SMAD) is the kinase whose direct substrate of TGF-beta receptor (SMAD1,2,3,5,8); (2) common type SMAD, its in the signal transduction cascade with R-SMAD effect (SMAD4); (3) inhibition type SMAD (SMAD6,7), it suppresses the proteic activity of above-mentioned SMAD.Among the various R-SMAD, SMAD2 and SMAD3 are TGF-β specific signals mediated factors, and in the cascade of TGF-signal, SMAD2/SMAD3 is enabled to interact with SMAD4 by I type TGF-beta receptor phosphorylation.The complex body of the SMAD2/SMAD3 of gained and SMAD4 can be indexed in the nucleus then, directly or by transcribing of other albumen initiation TGF-β regulatory gene (is summarized in Itoh (2000) Eru J Biochem 267:6954-6967 here; Shi (2003) Cell 113:685-700).

The function spectrum of TGF-β is very wide, and depends on cell type and differentiation state (Roberts (1990) Hand-book of Experimental Pharmacology:419-472).The cell function that TGF-β can influence comprises: apoptosis, propagation, differentiation, movability and cell adhesion.Therefore, TGF-β plays a significant role in multiple different bioprocess.During fetal development, TGF-β forms the site in form, particularly have the interactional zone of epithelial cell-mesenchymal cell expresses and induces important atomization (Pelton (1991) J Cell Biol 115:1091-1105).TGF-β also brings into play keying action (Mishra (2005) Science 310:68-71) at the self of undifferentiated state stem cell with in keeping.In addition, TGF-β also plays a significant role in immune regulation and control.It has immunosuppressive action usually, because it can suppress lymphopoiesis and limit the activity of tissue macrophages.Therefore thereby TGF-β can make inflammatory reaction disappear once more and help prevention over-drastic immune response (Bogdan (1993) Ann NY Acad Sci 685:713-739 is summarized in Letterio (1998) Annu RevImmunol 16:137-161).Another function of TGF-β is a regulating cell propagation.TGF-β suppresses the growth of endotheliocyte, epithelial cell and hematopoietic cell, but promote growth (Tucker (1984) the Science 226:705-707 of mesenchymal cell, Shipley (1986) Cancer Res 46:2068-2071, Shipley (1985) PNAS 82:4147-4151).Another critical function of TGF-β is that regulating cell adheres to and cell-cell interaction.For example Fiberonectin and collagen protein promote the accumulation of extracellular matrix to TGF-β by inducing extracellular matrix protein.In addition, TGF-β reduces the metalloprotease of matrix degradation and expression (Roberts (1990) the Ann NY Acad Sci 580:225-232 of inhibitors of metalloproteinase; Ignotz (1986) J Biol Chem 261:4337-4345; Overall (1989) J BiolChem 264:1860-1869, Edwards (1987) EMBO J 6:1899-1904).

The extensive sphere of action of TGF-β means TGF-β, and for example wound healing and pathologic process for example play a significant role in cancer and the fibrosis in a lot of physiological statuss.TGF-β is one of key somatomedin in the wound healing (being summarized in O ' Kane (1997) Int J Biochem Cell Biol 29:79-89).Form the phase in granulation tissue, TGF-β discharges from thrombocyte in injury site, and regulation and control generate himself in scavenger cell then, and for example induces monocyte to secrete other somatomedin.Most important function comprises the chemotaxis that stimulates inflammatory cell in the wound healing, and synthetic extracellular matrix is also regulated and control propagation, differentiation and the genetic expression of all important cells types that relate in the wound healing process.

Under pathological conditions, the beta mediated effect of these TGF-, particularly regulate and control the effect that extracellular matrix (ECM) generates and to cause fibrosis or skin scar (being summarized in Border (1994) N Engl J Med331:1286-1292).

For fibrotic conditions, diabetic nephropathy and glomerulonephritis, shown that TGF-β promotes nephrocyte pathologic loose and extracellular matrix to accumulate.By preventing the expansion of glomerular mesangium matrix, carrying out property of renal function weakens and in diabetic animal, reduce number (Border (1990) 346:371-374 that has formed damage in the diabetic renal glomerulus ephrosis with anti-TGF-beta antibodies handling interrupt TGF-signal transduction path, Yu (2004) Kindney Int 66:1774-1784, Fukasawah (2004) Kindney Int 65:63-74, Sharma (1996) Diabetes 45:522-530).

TGF-β also plays a significant role in hepatic fibrosis.TGF-β stimulates the activation that develops necessary stellate cells for hepatic fibrosis to generate myofibroblast, and forming the myofiber cell is the main generation person of extracellular matrix in the liver cirrhosis evolution.Similarly, shown that interruption TGF-signal transduction path can reduce fibrosis (Yata (2002) the Hepatology 35:1022-1030 in the experimental model; Aria (2003) BMC Gastroenterol 3:29).TGF-β also brings into play key function and (is summarized in Derynck (2001) Nature Genetics:29:117-129 in the forming process of cancer; Elliott (2005) J Clin Onc 23:2078-2093).Early stage in cancer development, TGF-β antagonism cancer forms, and this tumor inhibition effect mainly suppresses epithelial cell splitted ability based on TGF-β.On the contrary, at advanced tumor, TGF-β promotes the formation of growth of cancers and metastasis, and it is attributable to the growth-inhibiting effect that most epithelial tumors develop into TGF-β and produces tolerance, and TGF-β also supports the growth of cancer cells simultaneously by other mechanism.These mechanism comprise and promote that blood vessel takes place, immunosuppressive action, and it promotes growth of tumour cell and avoids immune controlled function (immunosurveillance) and the formation of promotion metastasis and strengthen its aggressive.The formation of tumor cell invasion prototype is the main prerequisite that metastasis forms.TGF-β promotes this process by the ability that its regulating cell adhesion, mobility and extracellular matrix form.In addition, the epithelium prototype of the beta induced cell of TGF-matter prototype between aggressive changes (EMT).In some researchs, the closely related property between TGF-β expression and the prognosis mala also can prove the vital role of TGF-β in promoting growth of cancers.Especially in prostate cancer, mammary cancer, intestinal cancer and patients with lung cancer, find TGF-β level rising (Wikstrom (1998) Prostate 37:19-29; Hasegawa (2001) Cancer 91:964-971; Friedman (1995) CancerEpidemiol Biomarkers Prev 4:549-54).

Because above-mentioned TGF-β is to the promoter action of cancer, it is a kind of possible treatment notion that inhibition TGF-signal transduction path for example suppresses this path by inhibition TGF-β I receptor.A lot of clinical preceding experiments have shown that interrupting TGF-signal transduction path suppresses growth of cancers really.Therefore, reduced the formation (Muraoka (2002) J Clin Invest 109:1551-1559, Yang (2002) J Clin Invest 109:1607-1615) of metastasis in the transgenic mice that develops into aggressive mammary cancer in time with the treatment of soluble T GF-β II receptor.

The tumor cell line of expressing defective TGF-β II receptor shows tumour and metastasis growth (Oft (1998) the Curr Biol 8:1243-1252 that weakens, McEachern (2001) Int J Cancer 91:76-82, Yin (1999) J Clin Invest 103:197-206).

" it is characterized by TGF-'beta ' activity strengthen " thus situation comprise TGF-β wherein synthetic be upset cause situation that TGF-β level raises wherein TGF-β latent albumen activated undesirably or be converted into the proteic situation of active TGF-β or wherein TGF-beta receptor situation about raising or wherein TGF-β albumen in the bonding force enhanced situation of disease location and cell or extracellular matrix.Therefore, in each case, " increased activity " is meant that all the biologic activity of the TGF-β that owing to what reason causes no matter arrives the high-caliber situation of not expecting.

Found that multiple disease generates too much relevant with TGF-β.

The inhibitor in TGF-β intracellular signal transduction path can be used for treating the fiber proliferative disease.Particularly, the fiber proliferative disease comprises and not modulated TGF-'beta ' activity and the excessive relevant kidney illness of fibrosis, comprises glomerulonephritis (GN), for example glomerular mesangium proliferative GN, immune GN, crescent GN.Other kidney illness comprises the transplant patient's of diabetic nephropathy, kidney region fibrosis, acceptance ring spore mattress element renal fibrosis and HIV associated kidney disease.Collagen vascular disease disease comprises progressive systemic sclerosis, polymyositis, the hardening of tissue, dermatomyositis, eosinophilic fasciitis, local scleroderma or the illness relevant with the morbidity of Raynaud syndrome.The pulmonary fibrosis that excessively causes by the TGF-'beta ' activity comprise adult respiratory distress syndrome, idiopathic pulmonary fibrosis with usually with autoimmune disorder for example systemic lupus erythematous and local scleroderma, chemistry contacts or allergy is relevant interstitial pulmonary fibrosis.Another autoimmune disease with fiber proliferative feature is a rheumatoid arthritis.

Drainage excessively generated relevant with TGF-β after the illness in eye relevant with the fiber proliferative disorders comprised the reattachment of retina of following the proliferative vitreoretinopathy, the cataract extraction of following the intra-ocular artificial lens implantation, glaucoma.

Generate excessively relevant fibrotic disease with TGF-β 1 and can be divided into for example for example corium scarization and restenosis (Chamberlain of the fibrosis of kidney, lung, liver and acute disease of chronic disease, J.Cardiovascular Drug Reviews, 19 (4): 329-344).Tumour cell also can cause immunosuppression to the synthetic and secretion of TGF-β 1, in the patient who suffers from aggressive brain tumor or mastadenoma visible (Arteaga etc., (1993) J.Clin.Invest.92:2569-2576).TGF-β 1 can acutely change the process (Barral-Netto etc., (1992) Science 257:545-547) of sharp ten graceful protozoan infection mouse.TGF-β 1 makes disease progression, and the TGF-beta 1 antibodies has then stopped the progress of disease in the mouse of susceptible in heredity.The mouse of tolerance becomes after using TGF-β 1 to sharp ten graceful protozoan infection susceptibles in the heredity.

About the summary of the profound influence of 1 pair of extracellular matrix deposition of TGF-β is seen (Rocco and Ziyadeh (1991), Contemporary Issues in Nephrology v.23, Hormones, autocoidsand the kidney.Ed.Jay Stein, Churchill Livingston, New York pp.391-410; Roberts etc., (1998) Rec.Prog.Hormone Res.44:157-197), its influence comprises stimulates extracellular matrix component to synthesize and suppress its degraded.Because the structure of renal glomerulus and filter character and form decision by the extracellular matrix of glomerular mesangium and mesangium to a great extent, thus 1 pair of kidney of TGF-β to have the profound influence this point not astonishing.At proliferative glomerulonephritis (Border etc., Kidney Int.37:689-695) and diabetic nephropathy (Mauer etc. (1990), (1984) J.Clin.Invest.74:1143-1155) accumulation of mesonephric glomerulus mesentery matrix also is the main pathological characteristics of described disease clearly.TGF-β 1 level rising (Yamamoto etc., (1993) Proc.Natl.Acad.Sci.90:1814-1818) in people's diabetic glomerulosclerosis (DPN late period).TGF-β 1 is important mediated factor (Phan etc., (1990) Kidney Int.37:426 in the pathogenic process at renal fibrosis in multiple Mammals model; Okuda etc., (1990) J.Clin.Invest.86:453).Antiserum(antisera) (Border etc. at TGF-β 1, (1990) Nature 346:371) and can be with TGF-β 1 bonded extracellular matrix protein, be that decorin (Border etc., (1992) Nature 360:361-363) shows and can suppress to test inductive rat glomerulonephritis.

Over-drastic TGF-β 1 causes the corium scar tissue to form.TGF-β 1 neutralizing antibody that is injected to rat healing wound edge has shown the inhibition cicatrization and has not disturbed speed of wound healing or wound tensile strength (Shah etc., (1992) Lancet 339:213-214).Blood vessel generation, scavenger cell and the monocyte number of wound all descend simultaneously, and destructive collegen filament deposition also descends in the scar tissue.

Behind balloon angioplasty, because one of factor that the carrying out property of arterial wall that smooth muscle cell proliferation and intra-arterial extracellular matrix deposition cause thickens may be TGF-β 1.The artery diameter of restenosis can be reduced 90% by this thickening, because the minimizing major part of diameter so may make these vessel open to 50% by reducing extracellular matrix sedimentation widely simply because extracellular matrix rather than smooth muscle cell body cause.In vivo transfection in the unmarred pig artery of TGF-β 1 gene, the genetic expression of TGF-β 1 and extracellular matrix synthetic relevant with hyperplasia (Nabel etc., (1993) Proc.Natl.Acad.Sci.USA 90:10795-10763).TGF-β 1 inductive hyperplasia does not resemble the PDGF-BB inductive extensive, but it is more extensive to contain the extracellular matrix of TGF-β 1 transfectant.In this transgenic pig model, extracellular matrix deposition and FGF-1 (a kind of secretor type of FGF) inductive hyperplasia have nothing to do (Nabel (1993) Nature 362:844-846).

In the cancer of several types, the TGF-β 1 that tumour generates may be deleterious.Become after with the carrier transfection of expressing mouse TGF-β 1 tumorigenicity and transitivity of MATLyLu rat prostate cancer cells (Stainer and Barrack (1992) Mol.Endocrinol 6:15-25) and MCF-7 human breast cancer cell (Arteaga etc., (1993) Cell Growth and Differ.4:193-201) is stronger.Blood vessel generation, transfer and prognosis mala relevant (Wikstrom, P. etc., (1998) (the Prostate 37:19-29 in TGF-β 1 and human prostata cancer and cancer of the stomach late period; Saito, H. etc., (1999) Cancer 86:1455-1462).In mammary cancer, prognosis mala and the rising of TGF-β level relevant (Dickson etc., (1987) Proc.Natl.Acad.Sci.USA 84:837-841; Kasid etc., (1987) Cancer Res.47:5733-5738; Daly etc., (1990) J.Cell Biochem.43:199-211; Barrett-Lee etc., (1990) Br.J Cancer 61:612-617; King etc., (1989) J.Steroid Biochem.34:133-138; Welch etc., (1990) Proc.Natl.Acad.Sci.USA 87:7678-7682; Walker etc., (1992) Eur.J.Cancer 238:641-644), with tamoxifen treatment inductive TGF-β 1 (Butta etc., (1992) Cancer Res.52:4261-4264) with tamoxifen treatment mammary cancer failure relevant (Thompson etc., (1991) Br.J.Cancer 63:609-614).Anti-TGF-beta 1 antibody suppresses the growth (Arteaga etc., (1993) J.Clin.Invest.92:2569-2576) of MDA-231 human breast cancer cell in athymic mouse, uses this antibody and spleen natural killer cell activity and increases dependency is arranged.Descend with the Chinese hamster ovary celI NK of latent TGF-β 1 transfection is active, tumor growth is accelerated (Wallick etc., (1990) J.Exp.Med.172:1777-1784) in nude mice.Therefore, breast tumor excretory TGF-β may cause the internal secretion immunosuppression.TGF-β 1 plasma concentration height shows mammary cancer patients with terminal prognosis mala (Anscher etc., (1993) N.Engl.J.Med.328:1592-1598).Before high dose chemotherapy and autologous bone marrow transplantation in the circulation the high patient of TGF-β level to suffer from the risk of hepatic veno-occulusive disease (all patients' 15-50%, mortality ratio is up to 50%) and idiopathic interstitial pneumonia (all patients' 40-60%) higher.These results' implication is 1) TGF-β 1 blood plasma level raises and can be used for differentiating high-risk patient and 2) minimizing of TGF-β 1 can reduce the M ﹠ M that these are used for patient with breast cancer's common treatment.

A lot of malignant cells secretion transforming growth factor-betas (TGF-β), it is an immunosuppressor efficiently, shows that it may be that a kind of tumour is escaped the important mechanism that host immune monitors that TGF-β generates.Setting up the TGF-signal ruined white corpuscle subgroup of transduceing in band knurl host is a kind of potential means of immunotherapy for cancer.The TGF-signal ruined transgenic animal model of transduceing can be eradicated normally the lymph tumor EL4 of the overexpression TGF-β of lethality (Gorelik and Flavell, (2001) Nature Medicine 7 (10): 1118-1122) in the T cell.

TGF-β excretory downward modulation in tumour cell can recover host immune originality, and the T cell is to the insensitive acceleration and the autoimmunization of breaking up of then causing of TGF-β, and these factors are for needing for the tumour of self antigen is expressed in antagonism among the host of tolerance.The immunosuppressive action of TGF-β is also relevant with a subgroup among the AIDS patient, estimates its immune response level lower (Garba etc., J.Immunology (2002) 168:2247-2254) according to its CD4/CD8 cell counting among these patients.TGF-β neutralizing antibody can reverse this effect in culture, show that TGF-signal transduction inhibitor can be used for reversing the AIDS patient's of this subgroup immunosuppression.

Take place in early days in cancer, TGF-β 1 can be used as tumor suppression efficiently, and can mediate the effect of some chemical protective agents.But, the growth of pernicious newborn knurl and in the progression certain a bit, as if tumour cell can be escaped TGF-β dependency growth-inhibiting, occurs biological activity TGF-β simultaneously in microenvironment.In the transgenosis system of overexpression TGF-β, the promoted dual function of this tumor suppression/tumour of TGF-β has obtained clarification the most clearly in eukaryotic cell.Although the transgenosis system tolerates the formation of optimum skin injury more, transfer conversion rate wherein but increases (Cui etc., (1996) Cell 86 (4): 531-42) greatly.As if the TGF-β 1 that malignant cell generates in the primary tumor increase along with the progress of neoplasm staging.In multiple main epithelial cancer, carry out studies show that human cancer to being increased in of TGF-β growing amount occur in the tumour progression process later relatively.In addition, this TGF-β relevant with tumour provides the selectivity advantage and promoted tumor growth for tumour cell.Interactional influence between TGF-β 1 pair cell interphase interaction and a cell and the matter can cause it to have higher invasion and attack and metastasis tendency.

Because the highly efficient depressor that the relevant TGF-β of tumour is the amplification of activated lymphocyte colony, so it can make tumour cell escape immunosurveillance.TGF-β also suppresses the generation of angiostatin.Cancer treatment method for example generates activated T GF-β in the radiation and chemotherapy induced tumor, thereby selects tolerant T GF-β Growth Inhibition malignant cell hypertrophy.Therefore, these anticancer therapies increase risk, make growth property and aggressive all be able to the accelerated development of enhanced tumour.In this case, the promoting agent in the beta mediated signal transduction path of target TGF-may be a kind of very effective therapeutic strategy.Tumour cell has shown the various kinds of cell toxic effect that can offset radiation and chemotherapy to the tolerance of TGF-β, between the treatment dependency activation of TGF-β may be more harmful in the matter, because TGF-β makes microenvironment be more conducive to tumor growth, may produce and cause Fibrotic tissue injury.Development TGF-signal transduction inhibitor uses separately or can be used in combination with other therapies all has the treatment of being beneficial to terminal cancer.

Described compound is suitable for treating cancer and other morbid state that influenced by TGF-β, and method is to suppress TGF-β among the patient who needs by described compound administration is being had in described patient.TGF-β also can be used for atherosclerosis (T.A.McCaffrey:TGF-ps and TGF-β Receptors in Atherosclerosis:Cytokine and Growth factor Reviews 2000,11,103-114) and alzheimer's disease (Masliah, E.; Ho, G.; Wyss-Coray, T.:Functional Role of TGF-β in Alzheimer ' s Disease Microvascular Injury:Lessons from Trangenic Mice:Neurochemistry International 2001,39,393-400).

Found that compound of the present invention and salt thereof have very valuable pharmacological character, tolerance is good simultaneously.

Particularly, it has TGF-beta receptor I kinase inhibition character.

Compound of the present invention preferably shows favourable biologic activity, and described biologic activity can easily prove in enzyme assay analytical method for example described herein.In this kind of enzyme analytical method, compound of the present invention preferably shows and causes restraining effect, usually can be by IC ₅₀Value proves in a suitable scope, is preferably micromole's level, more preferably in the nmole level.

As described herein, these signal transduction paths are relevant with multiple disease.Therefore, compound of the present invention can be used for preventing and/or treating by interacting with one or more described signal transduction path and depends on the disease in described signal transduction path.

Therefore, the present invention relates to promotor or inhibitor, preferably as the compound of the present invention of inhibitor as signal transduction as herein described path.Therefore the present invention preferably relates to promotor or the inhibitor as TGF-signal transduction path, preferably as the compound of the present invention of inhibitor.

The invention still further relates to one or more compounds of the present invention and be used for the treatment of and/or prophylactic purposes, the described disease disease that causes, mediates and/or propagate that increases by the TGF-'beta ' activity preferably described herein.

Therefore the present invention relates to be used for the treatment of and/or prevent the compound of the present invention of described disease, compound of the present invention to be used to prepare the purposes that is used for the treatment of and/or prevents the medicine of described disease as medicine and/or active constituents of medicine, and the method that is used for the treatment of described disease, it comprises one or more compound administration of the present invention in the patient of this treatment of needs.

Host or patient can belong to any mammalian species, for example primates, particularly human; Rodents comprises mouse, rat and hamster; Rabbit, horse, milk cow, dog, cat etc.Animal model can be used for experimental study, for the treatment human diseases supplies a model.

Concrete cell can be determined by in vitro tests for the susceptibility of The compounds of this invention.Usually cell culture is mixed one period that is enough to make the promoting agent inducing cell death or suppresses migration with the The compounds of this invention of various different concns, be generally about 1 hour to 1 week.Experiment in vitro can use the biopsy samples cultured cells.Count after handling still survival cells order then.

According to the compound of concrete use, disease specific, patient's states etc., dosage can change.Therapeutic dose is enough to significantly reduce the quantity of not expecting cell in the target tissue usually, keeps patient's viability simultaneously.Treatment continues to usually and cell burden has taken place significantly reduces, and for example the cell burden reduces at least about 50%, and can continue to and detect in vivo basically less than till the cell of not expecting.

For identification signal transduction path and detect interaction between each signal transduction path, a lot of scientists have developed proper model or model system, cell culture model (Khwaja etc. for example, EMBO, 1997 for example, 16,2783-93) and transgenic animal model (White etc. for example, Oncogene, 2001,20,7064-7072).Be to determine some stage of signal transduction cascade, can use interactional compound with conditioning signal (Stephens etc. for example, Biochemical J., 2000,351,95-105).Compound of the present invention also can be used as reagent and is used in animal and/or cell culture model or test kinase dependent signals transduction path in the described clinical disease of the application.

The measurement of kinase activity is a technology well known to those skilled in the art.The use substrate that is used to measure kinase activity is histone (Alessi etc. for example for example, FEBS Lett.1996,399,3, pp333-338) or the hereditary test macro of alkaline myelin protein be described in the document (Campos-Gonzalez for example, R. and Glenney, Jr., J.R.1992, J.Biol.Chem.267, pp 14535).

For differentiating kinase inhibitor, can utilize various analytical systems.The flicker proximate analysis (Sorg etc., J.of Biomolecular Screening, 2002,7,11-19) and during Flashplate analyzes, measure protein or peptide substrates by γ ATP radiophosphorus acidifying level.In the presence of the inhibition compound, can detect that radiated signal weakens or detected at all less than radiated signal.In addition, suitable analytical procedure also have the homogeneous phase time discrimination fluorescence resonance energy shift (HTR-FRET) and fluorescence polarization (FP) technology (Sills etc., J.of Biomolecular Screening, 2002,191-214).

Other on-radiation elisa assay method is used specific phosphoric acid antibody (phosphoric acid-AB).Phosphoric acid-AB only combines with the substrate of phosphorylation.Use the peroxidase-conjugated anti-goat antibody of secondary to detect this combination (Ross etc., 2002, Biochem.J. is about to deliver, manuscript BJ20020786) by chemoluminescence method.

Prior art

DE 2 409 308 has described compound " B2 ", " B3 ", " B4 ", " B5 ", " B8 ", " B9 ", " B10 ", " B12 ", " B14 ", " B16 ", " B17 ", " B19 ", " B20 " as active constituents of medicine, and it has analgesia and/or inflammatory effects.

E.Szarvasi etc. are at Eur.J.Med.1978,13, described compound " B1 ", " B6 ", " B7 ", " B11 ", " B13 ", " B15 ", " B18 ", " B21 ", " B22 " as active constituents of medicine among the 113-119, it has analgesia and/or inflammatory effects.

Other triazolo-1 has been described, the 5-benzodiazepine among the DE 2 318 673 Compounds.

L.Kosychova etc. are at Chemistry of Heterocyclic Compounds, and Vol.40 has described other and has been used to resist 5 of tumour, 6-dihydro-4H-1,2,4-triazolo-a among the 811-815 (2004)]-1,5 benzodiazepine Compound.

V.Ambrogi etc. are at II Farmaco 48, described among the 665-676 (1993) central nervous system is had active 1,4-benzothiazine and 1,5-benzothiazepine

Compounds.

Other triazole derivative as the TGF-beta inhibitor is disclosed among WO 03/042211 A1.

Other triazole derivative as the TGF-beta inhibitor has also been described among WO 2004/026307 A1.

Two cyclopyrrole derivatives as the TGF-beta inhibitor have been described among the WO 02/094833.

Summary of the invention

The present invention relates to be selected from down the compound of group:

And pharmaceutically acceptable derivative, solvate, salt, tautomer and steric isomer, comprise the mixture of their various ratios.

The invention still further relates to optically active isomer (steric isomer), enantiomorph, racemic modification, diastereomer and the hydrate and the solvate of these compounds.The solvate of term compound is meant that the inert solvent molecule adds and is bonded in the compound, owing to mutual magnetism forms.For example solvate has monohydrate or dihydrate or alcoholate.

The pharmaceutically acceptable derivative of term is meant for example salt and the so-called preceding drug compound of The compounds of this invention.

The term prodrug derivant is meant the compound of modifying by for example alkyl or carboxyl groups, sugar or oligopeptides of the present invention, and it is formed active compound of the present invention by rapid cracking in vivo.These also comprise the biodegradable polymer derivant of The compounds of this invention, Int.J.Pharm.115 for example, and 61-67 (1995) is described.

Representation " significant quantity " is meant that medicine or pharmacy activity component cause for example investigator or doctor's pursuit or the biology of expectation or the amount of medicinal response in tissue, system, animal or human's class.In addition, representation " treatment significant quantity " is meant with the corresponding individuality of not accepting this amount and compares, produces following result's amount: improve treat, heal, prevent or eliminate a disease, symptom, illness, main suit, symptom or side effect or slow down the progress of disease, main suit or illness simultaneously.

Representation " treatment significant quantity " also comprises the amount that can effectively strengthen normal physiological function.

The invention still further relates to the purposes of the mixture of The compounds of this invention, the mixture of two kinds of diastereomers for example, for example its ratio is 1: 1,1: 2,1: 3,1: 4,1: 5,1: 10 or 1: 1000.The mixture of preferred especially steric isomer compound.

Prepare compound of the present invention and be used to prepare their raw material according to known method own, (tool master book Houben-Weyl for example for example described in document, Methoden derorganischen Chemie[organic chemistry method], Georg-Thieme-Verlag, Stuttgart), accurately carry out under the known conditions of described reaction being suitable for.Can also employing itself known but this paper version in greater detail not.

Preferably by making tetrahydro benzo [b] [1,4] diaza

-2-40 thione derivatives and carbohydrazide compounds prepared in reaction compound of the present invention.

Reaction is carried out in inert solvent usually.The condition that depends on use, the reaction times day is not waited from several minutes to 14, and temperature of reaction is generally 30-130 ℃, particularly 60-120 ℃ for-15 to 150 ℃ approximately.

Suitable inert solvent for example has hydro carbons for example hexane, sherwood oil, benzene, toluene or dimethylbenzene; Chlorinated hydrocarbon is trieline, 1 for example, 2-ethylene dichloride, tetracol phenixin, chloroform or methylene dichloride; Alcohols is methyl alcohol, ethanol, Virahol, n-propyl alcohol, propyl carbinol or the trimethyl carbinol for example; Ethers is diethyl ether, diisopropyl ether, tetrahydrofuran (THF) (THF) or dioxan for example; Glycol ether is methyl glycol or an ether, glycol dimethyl ether for example; Ketone is acetone or butanone for example; Amides is ethanamide, N,N-DIMETHYLACETAMIDE or dimethyl formamide (DMF) for example; Nitrile is acetonitrile for example; Sulfoxide is methyl-sulphoxide (DMSO) for example; Dithiocarbonic anhydride; Carboxylic acid is formic acid or acetate for example; Nitro-compound is Nitromethane 99Min. or oil of mirbane for example; The ester class is the mixture of ethyl acetate or described solvent for example.

Preferred especially 1-butanols.

Handle under standard conditions with boron tribromide and to be suitable for the cracking ethers.

Pharmaceutical salts and other form

Described compound of the present invention can be used with their final salt-independent shapes.On the other hand, the present invention also comprises the purposes of these compounds of pharmaceutical acceptable salt, and described salt can be by methods known in the art derived from multiple organic and inorganic bronsted lowry acids and bases bronsted lowry.The pharmaceutical acceptable salt of most of The compounds of this invention can prepare by ordinary method.If The compounds of this invention comprises carboxylic group, one of its suitable salt can form by this compound and suitable alkali reaction are obtained corresponding base addition salt.This type of alkali is for example alkali metal hydroxide, comprises potassium hydroxide, sodium hydroxide and lithium hydroxide; Alkaline earth metal hydroxides is as hydrated barta and calcium hydroxide; Alkali metal alcoholates, for example potassium ethylate and sodium propylate; And multiple organic bases, as piperidines, diethanolamine and N-methyl glutamine.The aluminium salt that also comprises The compounds of this invention.In the situation of some The compounds of this invention, acid salt can be by forming these compounds with pharmaceutically useful organic and mineral acid treatment, and described acid is haloid acid for example, example hydrochloric acid, Hydrogen bromide or hydroiodic acid HI; Other mineral acid and their corresponding salt are as vitriol, nitrate or phosphoric acid salt etc.; And alkyl-and single arylsulphonate, as esilate, tosylate and benzene sulfonate; And other organic acid and their corresponding salt, as acetate, trifluoroacetate, tartrate, maleate, succinate, Citrate trianion, benzoate, salicylate, ascorbate salt etc.Therefore, the pharmaceutically acceptable acid additive salt of The compounds of this invention comprises following: acetate, adipate, alginate, arginic acid salt, aspartate, benzoate, benzene sulfonate, hydrosulfate, hydrosulphite, bromide, butyrates, camphorate, camsilate, octylate, muriate, chloro benzoate, Citrate trianion, cyclopentane propionate, digluconate, dihydrogen phosphate, dinitro-benzoate, dodecyl sulfate, esilate, fumarate, semi-lactosi hydrochlorate (deriving from tetrahydroxyadipic acid), the galacturonic hydrochlorate, gluceptate, gluconate, glutaminate, glycerophosphate, hemisuccinic acid salt, Hemisulphate, enanthate, hexanoate, hippurate, hydrochloride, hydrobromate, hydriodate, the 2-isethionate, iodide, isethionate, isobutyrate, lactic acid salt, Lactobionate, malate, maleate, malonate, mandelate, metaphosphate, mesylate, tolyl acid salt, monohydric phosphate, the 2-naphthalenesulfonate, nicotinate, nitrate, oxalate, oleate, palmitate, pectate, persulphate, phenylacetic acid salt, 3-phenylpropionic acid salt, phosphoric acid salt, phosphonate, phthalate, but the invention is not restricted to these salt.

In addition, the alkali salt of The compounds of this invention comprises aluminium, ammonium, calcium, copper, iron (III), iron (II), lithium, magnesium, manganese (III), manganese (II), potassium, sodium and zinc salt, but the invention is not restricted to these salt.Salt for mentioned above is preferably ammonium salt; The salt of alkali metallic sodium and potassium, and the salt of alkaline earth metals calcium and magnesium.Salt derived from the The compounds of this invention of pharmaceutically useful organic nontoxic alkali comprises primary amine, the salt of secondary amine and tertiary amine, the salt of substituted amine, also comprise naturally occurring substituted amine, the salt of the ion exchange resin of cyclammonium and alkalescence, arginine for example, trimethyl-glycine, caffeine, chloroprocaine, choline, N, N '-dibenzyl-ethylenediamin (benzyl star), dicyclohexylamine, diethanolamine, diethylamine, the 2-DEAE diethylaminoethanol, the 2-dimethylaminoethanol, thanomin, quadrol, N-ethylmorpholine, N-ethylpiperidine, glycosamine, glucosamine, Histidine, Hai Baming (hydrabamine), isopropylamine, lignocaine, Methionin, meglumine, N-methyl D-glycosamine, morpholine, piperazine, piperidines, versamid 900, PROCAINE HCL, PHARMA GRADE, purine, Theobromine, trolamine, triethylamine, Trimethylamine 99, the salt of tripropyl amine and trihydroxymethylaminomethane (tromethane), but this is not limited to the present invention.

The compound of the present invention that comprises alkaline nitrogen-containing group can be quaternized with it with following material: (C ₁-C ₄) alkyl halide, for example methyl, ethyl, sec.-propyl and t butyl chloride, bromide and iodide; Two (C ₁-C ₄) alkyl sulfuric ester, for example dimethyl, diethyl and diamyl sulfuric ester; (C ₁₀-C ₁₈) alkyl halide, for example decyl, dodecyl, lauryl, myristyl and octadecyl chlorination thing, bromide and iodide; And aryl (C ₁-C ₄) alkyl halide, for example benzyl muriate and styroyl bromination thing.Water-soluble and oil-soluble compound of the present invention all can use this type of salt preparation.

Preferred pharmaceutical salts mentioned above comprises acetate, trifluoroacetate, benzene sulfonate, Citrate trianion, fumarate, gluconate, hemisuccinic acid salt, hippurate, hydrochloride, hydrobromate, isethionate, mandelate, meglumine, nitrate, oleate, phosphonate, Pivalate, sodium phosphate, stearate, vitriol, sulfosalicylate, tartrate, Thiomalate, tosylate and tromethane, but this is not limited to the present invention.

The acid salt of the The compounds of this invention of alkalescence can form salt by the required acid-respons with free alkali form and capacity with ordinary method.Free alkali can be regenerated by this salt form is separated with alkali reaction and with free alkali with ordinary method.Free alkali form is different from their corresponding salt forms in some aspects, as some physical properties (for example solubleness in polar solvent); Yet with regard to purpose of the present invention, salt and they free alkali form separately is suitable.

As mentioned before, the base addition salt of pharmaceutically useful The compounds of this invention can be formed by metal or amine, for example basic metal and alkaline-earth metal or organic amine.Preferred metal is sodium, potassium, magnesium and calcium.Preferred organic amine is N, N '-dibenzyl-ethylenediamin, chloroprocaine, choline, diethanolamine, quadrol, N-methyl D-glycosamine and PROCAINE HCL, PHARMA GRADE.

The base addition salt of tart compound of the present invention can form salt by the required alkali reaction with free acid form and capacity with ordinary method.Free acid can be regenerated by this salt form is separated with acid-respons and with free acid with ordinary method.Free acid form is different from their corresponding salt forms in some aspects, as some physical properties (for example solubleness in polar solvent); Yet with regard to purpose of the present invention, salt and they free acid form separately is suitable.

If compound of the present invention comprises the group that can form this type of pharmacologically acceptable salt more than, the present invention also comprises many salt so.Typical many salt forms comprise for example two tartrates, diacetin, two fumarates, two meglumines, diphosphate, disodium salt and tri hydrochloride, but this is not limited to the present invention.

As indicated above, term herein " pharmacologically acceptable salt " is meant the activeconstituents of the The compounds of this invention that comprises a kind of salt form as can be known, if particularly this salt form is compared when making moderate progress the free form of the pharmacokinetic property of activeconstituents and the activeconstituents that uses before or any other salt form of activeconstituents.The required pharmacokinetic property that the pharmaceutical acceptable salt of activeconstituents can not have before providing first for this activeconstituents, and even can have positive effect about the pharmacodynamics of its cylinder therapeutic effect aspect to this activeconstituents.

In addition, the present invention relates to comprise at least a The compounds of this invention and/or its pharmaceutically useful derivative, solvate and steric isomer, comprise the mixture of its all proportions, and the optional vehicle and/or the medicine of assistant agent.

Pharmaceutical preparation can be with the form administration of dose unit, and this dose unit comprises the activeconstituents of every dose unit predetermined amount.This dose unit can comprise for example The compounds of this invention of 0.5mg-1g, preferred 1mg-700mg, preferred especially 5mg-100mg, concrete dosage depends on disease, administering mode and patient's age, body weight and the situation of being treated, perhaps pharmaceutical preparation can be with the form administration of dose unit, and this dose unit comprises the activeconstituents of every dose unit predetermined amount.The preferred dosage unit formulation is those preparations that comprise the activeconstituents of per daily dose mentioned above or broken dose or its corresponding section.In addition, this type of pharmaceutical preparation can be used the known method of pharmaceutical field and prepares.

Pharmaceutical preparation can be by any required proper method administration, for example by oral (comprising oral cavity or hypogloeeis), rectum, nose, part (comprise oral cavity, hypogloeeis or through skin), vagina or non-stomach and intestine (comprising subcutaneous, intramuscular, intravenously or intradermal) mode administration.This type of preparation can be used the known method of all pharmaceutical fields and prepare by for example activeconstituents being mixed with vehicle or assistant agent.

The pharmaceutical preparation that is applicable to oral administration can be with the unit administration that separates, as capsule or tablet; Powder or granule; Solution in water or on-aqueous liquid or suspensoid; Edible foaming agent or foam food; Perhaps oil-in-water liquid emulsion or water-in-oil liquid emulsion.

Therefore, for example, under with tablet or Capsule form case of oral administration, activeconstituents and oral, nontoxic and pharmaceutically useful inert excipient such as ethanol, glycerine, water etc. can be mixed.Powder can prepare by compound powder being broken into suitable fine powder and it being mixed with the drug excipient of pulverizing with similar approach, and described vehicle is for example edible carbohydrate, as starch or N.F,USP MANNITOL.Also can use correctives, sanitas, dispersion agent and dyestuff etc.

Capsule can prepare by preparing aforesaid powdered mixture and it being inserted in the gelatin shell of shaping.The polyoxyethylene glycol of glidant and lubricant such as high dispersive silicic acid, talcum powder, Magnesium Stearate, calcium stearate or solid form can join in the powdered mixture before stuffing operation.Can add disintegrating agent or solubilizing agent for example agar, lime carbonate or yellow soda ash equally, so that after taking capsule, improve the availability of medicine.

In addition, if desired or necessary, also can in mixture, mix suitable adhesive, lubricant and disintegrating agent and dyestuff.Suitable adhesive comprises starch, gelatin, natural sugar (as glucose or beta lactose, with the sweeting agent of corn preparation), natural and synthetic rubber (as gum arabic, tragacanth gum or sodium alginate), carboxymethyl cellulose, polyoxyethylene glycol, wax etc.Used lubricant comprises sodium oleate, sodium stearate, Magnesium Stearate, Sodium Benzoate, sodium acetate, sodium-chlor etc. in these formulations.Disintegrating agent includes but not limited to starch, methylcellulose gum, agar, wilkinite, xanthan gum etc.Tablet is by for example preparing powdered mixture, granulation or dry method pressing mixt, adding lubricant and disintegrating agent and suppressing whole mixtures and prepare to obtain tablet.Powdered mixture can will mix with thinner or matrix with the compound that proper method is pulverized by as indicated above, and randomly with tackiness agent (for example carboxymethyl cellulose, alginates, gelatin or polyvinylpyrrolidone), dissolving retarding agent (as paraffin), absorb accelerator (as quaternary salt) and/or sorbent material (as wilkinite, kaolin or Lin Suanergai) and mix and prepare.Powdered mixture can by with tackiness agent (as the solution of syrup, starch slurry, mucialga of arabic gummy or Mierocrystalline cellulose or polymer materials) with it wetting and extruding sieve and granulate.As the replacement scheme of granulating, powdered mixture can be operated by tabletting machine, is pressed into the agglomerate that shape differs, and this agglomerate is pulverized the back and formed particle.Particle can lubricate by adding stearic acid, stearate, talcum powder or mineral oil, adheres to tablet die to avoid it.Suppress lubricated mixture then to obtain tablet.Compound of the present invention also can mix with free-pouring inert excipient, obtains tablet without granulation or the direct compacting of dry method pressing step then.Can have transparent or opaque protective layer, this protective layer comprises lac sealing ply, sugar or polymer material layer and wax polishing layer.Can in these Drug coatings, add dyestuff so that distinguish different dose units.

Oral liquid is solution, syrup and elixir for example, can prepare with dosage unit form, so that it comprises the compound of predetermined amount.Syrup can prepare by compound is dissolved in the aqueous solution that contains suitable correctives, and elixir can be used the preparation of non-toxic alcohol medium.Suspensoid can prepare by compound is scattered in the nontoxic medium.Also can add solubilizing agent and emulsifying agent (as the pure and mild polyoxyethylene sorbitol ether of isooctadecane of ethoxylation), sanitas, flavoring additive (as spearmint oil or natural sweeting agent or asccharin) or other artificial sweetening agent etc.

If desired, the dosage unit preparations of oral administration can be packed in the micro-capsule.Said preparation also can prepare in the mode that prolongs or delay to discharge, and for example is embedded in polymkeric substance, the wax etc. by dressing or with microparticle material.

The compounds of this invention and salt thereof, solvate and derivative that physiologic function arranged also can be with the form administrations of liposome transfer system, for example individual layer vesicles, the big vesica of individual layer and multilamellar vesicle.Liposome can for example cholesterol, stearylamine or phosphatidylcholine form by various phosphatide.

The compounds of this invention and salt thereof, solvate and have the derivative of physiologic function can be with transmitting as single carrier with this compound molecule link coupled monoclonal antibody.Compound also can with the soluble polymer coupling as target medicine carrier.This base polymer can comprise polyvinylpyrrolidone, pyran co-polymer, poly-hydroxypropylmethyl acrylamido phenol, poly-hydroxyethyl aspartoyl amino-phenol or polyoxyethylene polylysine, and it can be replaced by palmitoyl.In addition, compound can also be fit to obtain the biodegradable polymkeric substance coupling of medicine controlled releasing with a class, and this polymkeric substance is the crosslinked or amphipathic segmented copolymer of poly(lactic acid), poly--6-caprolactone, polyhydroxybutyrate, poe, polyacetal, poly-dihydroxyl pyrans, polybutylcyanoacrylate and hydrogel for example.

The pharmaceutical preparation that is suitable for percutaneous dosing can be to be used for the long-time form administration that closely contacts the independent plaster of acceptor epidermis.Therefore, for example activeconstituents can pass through as Pharmaceutical Research from plaster, and 3 (6), the iontophoresis transmission described in 318 (1986).

The medicinal compound that is suitable for topical can be formulated as ointment, ointment, suspensoid, lotion, powder, solution, paste, gelifying agent, sprays, aerosol or finish.

For example for the treatment of oral cavity and skin, preparation is preferably used with the form of topical ointments or ointment for eye or other outside organization.At preparation is under the situation of ointment, and activeconstituents can be with paraffin or easily and the miscible ointment base common application of water.Perhaps, activeconstituents can be mixed with ointment with oil-in-water-type ointment matrix or water-in-oil-type ointment matrix.

The pharmaceutical preparation that is suitable for being applied topically to eye comprises eye drops, and wherein activeconstituents is dissolved in or is suspended in the suitable carrier, particularly in the aqueous solvent.

The pharmaceutical preparation that is suitable for being applied topically to the oral cavity comprises lozenge, pastille and mouth wash shua.

The pharmaceutical preparation that is suitable for rectal administration can be with the form administration of suppository or enema.

The pharmaceutical preparation (wherein carrier substance is a solid) that is suitable for intranasal administration comprises having for example dust base in the 20-500 micrometer range of particle diameter, and it is with the mode administration of snuffing, promptly by nasal meatus near sucking fast in the container that powder is housed of nasal cavity.With the water or the oil solution that are suitable for the preparation of the form administration of nasal spray or nasal drop comprise activeconstituents of liquid as carrier substance.

The pharmaceutical preparation that is suitable for inhalation comprises fine grain powder or spraying, and it can produce by polytype pressurized dispersion device with aerosol, atomizer or insufflator.

The pharmaceutical preparation that is suitable for vagina administration can be with vaginal suppository, vagina plug, ointment, gelifying agent, paste, foaming agent or spray agent administration.

The pharmaceutical preparation that is suitable for parenteral admin comprises water-based and the non-water aseptic parenteral solution that contains antioxidant, buffer reagent, presses down mattress agent and solute, and said preparation and the patient's who is treated blood etc. is opened; And water-based and non-water sterile suspension, it can comprise suspending medium and thickening material.These preparations can carry out administration with the container of single dose or multiple doses, for example Mi Feng ampoule and bottle, and be stored under lyophilize (freeze-drying) state, thereby only need face with before add sterile liquid carrier, water for injection for example.Injection solution and suspension according to the prescription preparation can be by no mattress powder, particle and tablet preparation.

Self-evident, except specifically noted component above, for particular type of formulation, also can comprise this area other material commonly used in the preparation; Therefore, the preparation that for example is suitable for oral administration can comprise correctives.

The treatment significant quantity of The compounds of this invention depends on multiple factor, comprises the character and the medication of the concrete illness of the age of animal for example and body weight, needs treatment and seriousness thereof, preparation, and is finally determined by doctor in charge or animal doctor.Yet compound of the present invention is used for the treatment of the significant quantity of tumor growth (for example colon or mammary cancer), is generally 0.1-100mg/kg acceptor every day (Mammals) body weight, particularly every day the 1-10mg/kg body weight.Therefore, for the Adult Mammals that body weight is 70kg, the actual amount of every day is generally 70-700mg, and wherein this dosage can be used as single dose administration every day, perhaps usually with a series of part dosage (as 2,3,4,5 or 6 times) administration every day, but total per daily dose is constant.Its salt or solvate or have the significant quantity of the derivative of physiologic function can be defined as the mark of the significant quantity of The compounds of this invention itself.Can infer the similar dosage that is suitable for above-mentioned other treatment of conditions.

The invention still further relates to and comprise at least a The compounds of this invention and/or its pharmaceutically useful derivative, solvate and steric isomer, comprise the mixture of its all proportions and the medicine of at least a other drug activeconstituents.

The present invention also relates to medicine box, this medicine box comprises independent packing

(a) The compounds of this invention of significant quantity and/or its pharmaceutically useful derivative, solvate and steric isomer comprise the mixture of its all proportions,

And

(b) the other drug activeconstituents of significant quantity.

Described medicine box comprises proper container, as box, independent bottle, bag or ampoule.This medicine box can comprise for example independent ampoule, and each ampoule comprises The compounds of this invention and/or its pharmaceutically useful derivative, solvate and the steric isomer of significant quantity, comprises the mixture of its all proportions,

And the other drug activeconstituents of the dissolving of significant quantity or lyophilized form.

Purposes

Be selected from down the compound of the present invention of group

And pharmaceutically acceptable derivative, salt, solvate, tautomer and steric isomer, the mixture that comprises their various ratios are suitable as active constituents of medicine and are used for Mammals, particularly human, the medicine that is used to prepare treatment and/or antagonism cancer, tumor growth, metastasis growth, fibrosis, restenosis, HIV infection, alzheimer's disease, atherosclerosis and/or promotes wound healing.

Preferred its is used for the treatment of the purposes that described disease is a solid tumor especially.

Described solid tumor is preferably selected from tesselated epithelium knurl, bladder cancer, cancer of the stomach, kidney, head and neck cancer, esophagus cancer, cervical cancer, thyroid carcinoma, intestinal cancer, liver cancer, the cancer of the brain, prostate cancer, genitourinary cancer, lymphsystem cancer, cancer of the stomach, laryngocarcinoma and/or lung cancer.

Solid tumor further is preferably selected from adenocarcinoma of lung, minicell adenocarcinoma of lung, carcinoma of the pancreas, glioblastoma, colorectal carcinoma and mammary cancer.

Further be preferred for treating the purposes of neoplastic hematologic disorder and immunity system tumour, preferred therapeutic is selected from the purposes of acute myelocytic leukemia, chronic granulocytic leukemia, kemia and/or chronic lymphatic leukemia.

Compound of the present invention also is suitable for and known carcinostatic agent combination.These known carcinostatic agents comprise and the following: estrogenic agents, androgen receptor modifier, retinoid receptor modulators, cytotoxic agent, antiproliferative, prenyl protein transferase inhibitor, HMG CoA-reductase inhibitors, hiv protease inhibitor, reverse transcriptase inhibitors and other angiogenesis inhibitor.Compound of the present invention is particularly suitable for using simultaneously with radiotherapy.Existing in the prior art describe (referring to the WO 00/61186) of synergy that suppresses VEGF and radiotherapy.

" estrogenic agents " is meant the compound that disturbs or suppress oestrogenic hormon and receptors bind with any mechanism.The example of estrogenic agents include but not limited to tamoxifen, raloxifene, idoxifene, LY353381, LY117081, toremifene, fluorine tie up this group, 4-[7-(2, the 2-dimethyl)-and 1-oxopropoxy-4-methyl-2-[4-[2-(piperidino)-oxyethyl group] phenyl]-2H-1-chromene-3-base-phenyl-2,2-dimethyl-propionic ester, 4,4 '-dihydroxy-benzene ketone-2,4-dinitrophenylhydrazone and SH646.

" androgen receptor modifier " is meant the compound that disturbs or suppress male sex hormone and receptors bind with any mechanism.The example of androgen receptor modifier comprises that Fei Nasi carries and its Mi Te of other 5 alpha reductase inhibitor, Nilutamide, fluorine, bicalutamide, liarozole, acetic acid Abiraterone.

" retinoid receptor modulators " is meant the compound that disturbs or suppress retinoid and receptors bind with any mechanism.The example of retinoid receptor modulators includes but not limited to that bexarotene, vitamin A acid, 13-are along vitamin A acid, 9-cis-retinoic acid, alpha-difluoromethyl ornithine, ILX23-7553, trans-N-(4 '-hydroxy phenyl) dimension methylamine and N-4-carboxyl phenyl-Wei methylamine.

" cytotoxic agent " is meant mainly by directly acting on cell function or inhibition or interference cell reduction division and causes the compound of necrocytosis, comprise alkylating agent, tumour necrosis factor, intercalator, microtubule inhibitor and topoisomerase enzyme inhibitor.

The example of cytotoxic agent includes but not limited to Win-59075; Sertenef; cachectin; ifosfamide; tasonermin; lonidamine; carboplatin; hexamethyl melamine; prednimustine; mitolactol; ranomustine; Fotemustine; S 254; oxaliplatin; Temozolomide; heptan platinum; estramustine phosphate; the toluenesulphonic acids improsulfan; Z-4828; Nidran; dibrospidium chloride; pumitepa; Lip river platinum; husky platinum; methylmitomycin; cis-platinum; irofulven; right ifosfamide; along amine dichloro (2-picoline) platinum; the benzyl guanine; glufosfamide; GPX100; (trans; trans; trans) two mu-(hexane-1; the 6-diamines)-mu-[diamines-platinum (II) two [diamines (chlorine) platinum (II)] tetrachloride; two aziridinyls-smart ammonia; ARSENIC TRI OXIDE 98; 1-(11-dodecyl amino-10-hydroxyl undecyl)-3, the 7-dimethyl xanthine; zorubicin; idarubicin; daunorubicin; Bisantrene; mitoxantrone; Perarubicin; pinafide; valrubicin; amrubicin; antineoplaston; 3 '-deaminizating-3 '-morpholinyl-13-deoxidation-10-hydroxyl Carubicin; Annamycin; galarubicin; Elinafide; MEN10755 and 4-demethoxylation-3-deaminizating-3-'-aziridino-4-methyl sulphonyl daunorubicin (referring to WO 00/50032).

The example of Antitubulin comprises taxol; vinealeucoblastine(VLB); 3 '; 4 '-two dehydrogenations-4 '-deoxidation-8 '-navelbine (norvincaleukoblastine); Docetaxel (docetaxol); rhizomycin; dolastatin; the mivobulin isethionate; Ao Ruitating (auristatin); Cemadotin; PRP109881; BMS184476; Vinflunine; Cryptophycin; 2; 3; 4; 5; 6-five fluoro-N-(3-fluoro-4-p-methoxy-phenyl) benzsulfamide; F 81097; N, N-dimethyl-L-is valyl-L-is valyl-and N-methyl-L-is valyl-L-prolyl-L-proline(Pro)-tert-butylamides; TDX258 and BMS188797.

Topoisomerase enzyme inhibitor has for example Hycamtin; Hycaptamine; Rinotecan; rubitecan; 6-ethoxy-c acyl group-3 '; the outer Ben Yajiaji chartreusin of 4 '-O-; 9-methoxyl group-N; N-dimethyl-5-nitropyrazole also [3; 4; 5-kl] acridine-2-(6H) propylamine; 1-amino-9-ethyl-5-fluoro-2; 3-dihydro-9-hydroxy-4-methyl-1H; 12H-benzo [de] pyrans also [3 '; 4 ': b; 7] indolizine also [1; 2b] quinoline-10; 13 (9H; 15H)-diketone; lurtotecan; 7-[2-(N-sec.-propyl amino) ethyl]-(20S) camptothecine; BNP1350; BNPI1100; BN80915; BN80942; the phosphoric acid Zuyeyidal; teniposide; sobuzoxane; 2 '-dimethylamino-2 '-deoxidation Zuyeyidal; GL331; N-[2-(dimethyl-amino) ethyl]-9-hydroxyl-5; 6-dimethyl-6H-pyrido [4; 3-b] carbazole-1-methane amide; Asulacrine; (5a; 5aB; 8aa; 9b)-9-[2-[N-[2-(dimethylamino) ethyl]-the N-methylamino] ethyl]-5-[4-hydroxyl-3; the 5-Dimethoxyphenyl]-5; 5a; 6; 8; 8a; 9-hexahydro furyl also (3 '; 4 ': 6; 7) naphthalene (2; 3-d)-1; 3-dioxo-6-ketone; 2; 3-(methylene-dioxy)-5-methyl-7-hydroxyl-8-methoxyl group benzo [c] phenanthridines; 6; two [(2-amino-ethyl) amino] benzo [g] isoquinoline 99.9-5 of 9-; the 10-diketone; 5-(3-amino propyl amino)-7; 10-dihydroxyl-2-(2-hydroxyethyl amino methyl)-6H-pyrazolo [4; 5; 1-de]-acridine-6-ketone; N-[1-[2-(diethylamino) ethylamino]-7-methoxyl group-9-oxo-9H-sulfo--xanthenes-4-ylmethyl] methane amide; N-(2-(dimethylamino) ethyl) acridine-4-methane amide; 6-[[2-(dimethylamino) ethyl] amino]-3-hydroxyl-7H-indeno [2,1-c] quinoline-7-ketone and dimesna.

" antiproliferative " comprises for example G3139 of sense-rna and DNA oligonucleotide; ODN698; RVASKRAS; GEM231 and INX3001 and antimetabolite be enocitabine for example; Mifurol; Ftorafur; pentostatin; doxifluridine; Trimetrexate; fludarabine; capecitabine; Galocitabine; cytosine arabinoside octadecyl sodium phosphate; fluorine is for shore sodium hydrate (fosteabine sodiumhydrate); Raltitrexed; Paltitrexid; emitefur; tiazofurine; Decitabine; Nolatrexed; pemetrexed; Nelzarabine; 2 '-deoxidation-2 '-methylene radical cytidine; 2 '-fluorine methylene radical-2 '-desoxycytidine; N-[5-(2; the 3-dihydro benzo furyl) alkylsulfonyl]-N '-(3; the 4-dichlorophenyl) urea; N6-[4-deoxidation-4-[N2-[2 (E); 4 (E)-14 carbon two enoyl-s] glycyl amino]-L-glycerine-B-L-pyrans mannoheptose base] VITAMIN B4; Aplidine; Ecteinascidin; troxacitabine; 4-[2-amino-4-oxo-4; 6; 7; 8-tetrahydrochysene-3H-Mi Dingbing [5; 4-b]-1; 4-triazine-6-base-(S)-ethyl]-2; 5-thiophene acyl group-L-L-glutamic acid; aminopterin-induced syndrome; 5 FU 5 fluorouracil; alanosine; 11-ethanoyl-8-(carbamoyloxy methyl)-4-formyl radical-6-methoxyl group-14-oxa--1; 11-diaza Fourth Ring-(7.4.1.0.0) 14-2; 4,6-triolefin-9-yl acetate; sphaerophysine; lometrexol; dexrazoxane; methioninase; 2 '-cyano group-2 '-'-deoxy-n 4-palmitoyl-1-B-D-arabinofuranosyl adenin cytosine(Cyt) and 3-aminopyridine-2-formaldehyde thiosemicarbazone.The monoclonal antibody that " antiproliferative " also comprises somatomedin of listing down except " angiogenesis inhibitor " is trastuzumab and tumor suppressor gene p53 for example for example, it can be sent (referring to for example United States Patent (USP) 6 by the transgenosis of recombinant virus mediation, 069,134).

Measure external (enzyme) analysis that inhibitor is renderd a service the inhibition of the beta mediated effect of TGF-

As an example, the test inhibitor is eliminated the beta mediated growth inhibiting ability of TGF-.

With pulmonary epithelial cells is that the Mv1Lu cell is inoculated on the 96 hole microtiter plates overnight incubation under standard conditions with the cell density of determining.Second day, substratum is replaced with the substratum that contains 0.5%FCS and 1ng/ml TGF-β, add determine concentration tried material, the normally form of 5 times of serial dilutions.The constant concentration of solvent DMSO is 0.5%.Two days later, with Viola crystallina with cell dyeing.From the fixed cell, extract after the Viola crystallina, in 550nm place spectrophotometer measurement optical density.It can be used for the adherent cell that quantitative assay exists, but the therefore propagation situation of cell in the quantitative culture process.

The inhibition of table 1:TGF-β

Compound number	IC 50[mol/l]
		“A1”	1.80E-07
“A2”	9.70E-07
		“A3”	3.50E-06
“A4”	3.80E-06
		“A5”	2.40E-06

“A6”	3.90E-06
		“A7”	2.20E-06
“A8”	6.60E-06
		“A9”	9.10E-06
“A10”	5.70E-06
		“A78”	1.5E-07
“A80”	3.3E-06
		“B1”	7.00E-06

The cell analysis of test TGF-β I receptor kinase inhibitor

The kinases analysis is carried out in 384 hole Flashplate.

With 31.2nM GST-ALK5,439nM GST-SMAD2 and 3mM ATP (0.3 μ Ci ³³The P-ATP/ hole) be to cultivate 45 minutes not containing or containing under 30 ℃ under the condition of being tried material (5-10 concentration) in the substratum (20mM HEPES, 10mM MgCl, 5mM MnCl, 1mM DTT, 0.1%BSA, pH7.4) of 35 μ l at cumulative volume.With 25 μ l200mM EDTA solution termination reactions, suction filtration at room temperature after 30 minutes.With 100 μ l 0.9%NaCl solution each hole is washed three times.In TopCount, measure radioactivity.Calculate IC with RS1 ₅₀Value.

Whole temperature in the context all are ℃.In following examples, " conventional aftertreatment " is meant: add entry when needed, when needed with pH regulator to 2-10, the composition that depends on end product, with ethyl acetate or dichloromethane extraction mixture, separate each phase, organic phase is also evaporated with dried over sodium sulfate, and product is through the silica gel chromatography purifying and/or through the crystallization process purifying.Rf value on the silica gel; Eluent: ethyl acetate/methanol 9: 1.

Mass spectrum (MS):

EI (electron bombardment ionization) M ⁺

FAB (fast atom bombardment) (M+H) ⁺

ESI (electronic spraying ionization) (M+H) ⁺

APCI-MS (atmospheric pressure chemical ionization mass spectrum) (M+H) ⁺

Retention time R _t[minute]: measure by HPLC

Post: Chromolith SpeedROD, 50x4.6mm ²(order number: 1.51450.0001 buys from Merck ﹠ Co., Inc.)

Gradient: 5.0 minutes, t=0 minute, A: B=95: 5, t=4.4 minute: A: B=25: 75, t=4.5 minute-5.0 minutes: A: B=0: 100

Flow velocity: 3.00ml/min

Eluent A: water+0.1%TFA (trifluoroacetic acid)

Eluent B: acetonitrile+0.08%TFA

Wavelength: 220nm

Embodiment 1

(R, S)-8-hydroxy-4-methyl-1-(6-picoline-2-yl)-5,6-dihydro-4H-2,3,6, the 10b-tetrazine also the preparation of [e] azulene (" A1 ") to carry out with the similar scheme of following scheme:

1.15-the preparation of methoxyl group-2-N-methyl-p-nitroaniline

10g 5-chloro-2-N-methyl-p-nitroaniline is dissolved in the 100ml methyl alcohol, adds the 32.3g sodium methylate.Reaction mixture was refluxed 18 hours under boiling.After the cooling, mixture is evaporated to dried, adds 500ml water, the filtering separation crude product.Obtain 9.15g 5-methoxyl group-2-N-methyl-p-nitroaniline after the drying.

1.2 (R, S)-preparation of 3-(5-methoxyl group-2-nitrophenyl amino)-2-methyl-propionic acid

9.15g 5-methoxyl group-2-N-methyl-p-nitroaniline is dissolved among the 60ml THF, adds 6.5ml isopropene cyanide and the solution of 1.35ml 40% benzyltrimethylammon.um oxyhydroxide in methyl alcohol.Reaction mixture was heated about 20 hours under boiling, and after the cooling, evaporation obtains the oily resistates.Crude product is dissolved in 400ml methyl alcohol and the 150ml water, adds 150ml 32% sodium hydroxide solution.With mixture heating 3 hours under boiling, cooling, evaporation, (R S)-3-(5-methoxyl group-2-nitrophenyl amino)-2-methyl-propionic acid, is crude product to obtain 8.9g through conventional aftertreatment.

1.3 (R, S)-8-methoxyl group-4-methyl isophthalic acid-(6-picoline-2-yl)-5,6-dihydro-4H-2,3,6, the 10b-tetrazine is the preparation of [e] azulene also

With the 8.9g of gained (R, S)-3-(5-methoxyl group-2-nitrophenyl amino)-2-methyl-propionic acid is dissolved in 40ml water and the 40ml acetate, adds 8.4g zinc (meal), with mixture heating 18 hours under boiling.Through the water-based aftertreatment obtain 2.35g (R, S)-7-methoxyl group-3-methyl isophthalic acid, 3,4,5-tetrahydro benzo [b]-1,4-diaza -2-ketone, its not purified subsequent reactions that promptly is used for.Crude product is dissolved in the 25ml pyridine, adds 5.1g 2, two (the 4-p-methoxy-phenyls)-2 of 4-, 4-dithio-1,3,2,4-dithia two phosphorus heterocycle butane heat mixture 3 hours down at 110 ℃.After the cooling, mixture is carried out the water-based aftertreatment, make the crude product precipitation by adding ether.Obtain after the drying 2.34g (R, S)-7-methoxyl group-3-methyl isophthalic acid, 3,4,5-tetrahydro benzo [b]-1,4-diaza

-2-thioketones.It was heated 17 hours at 110 ℃ in 5ml 1-butanols with 6-picoline-2-carbohydrazide.The cooling and after the water-based aftertreatment, with the oily crude product through the preparation HPLC purifying.Obtain 78mg (R, S)-8-methoxyl group-4-methyl isophthalic acid-(6-picoline-2-yl)-5,6-dihydro-4H-2,3,6, the 10b-tetrazine is [e] azulene also.

1.4 with 60mg (R, S)-8-methoxyl group-4-methyl isophthalic acid-(6-picoline-2-yl)-5,6-dihydro-4H-2,3,6, the 10b-tetrazine also [e] azulene is dissolved in the 2.5ml methylene dichloride, drips 162 μ l boron tribromides.After at room temperature reacting 6 hours, mixture is evaporated to dried, crude product obtains 40mg " A1 " through the preparation HPLC purifying.

Obtain following compound through similar approach:

Following examples relate to medicine:

Embodiment A: injection vials

With the hydrochloric acid of 2N activeconstituents and the solution of 5g Sodium phosphate dibasic in the 3L redistilled water of 100g formula I is adjusted to pH 6.5, sterile filtration is transferred in the injection vials, freeze-drying and in sealed under aseptic conditions under aseptic condition.Each injection vials contains the 5mg activeconstituents.

Embodiment B: suppository

The activeconstituents of 20g formula I and the mixture of 100g soybean lecithin and 1400g theobroma oil are melted, pour in the mould and make it and cool off.Each suppository contains the 20mg activeconstituents.

Embodiment C: solution

By with the activeconstituents of 1g formula I, the NaH of 9.38g ₂PO ₄2H ₂The Na of O, 28.48g ₂HPO ₄12H ₂The benzalkonium chloride of O and 0.1g is dissolved in the 940ml redistilled water and prepares solution.PH is transferred to 6.8, this solution is formulated into 1L and passes through radiation sterilization.This solution can be used as eye drop.

Embodiment D: ointment

Activeconstituents with 500mg formula I under aseptic condition mixes with 99.5g Vaseline.

Embodiment E: tablet

The mixture of activeconstituents, 4kg lactose, 1.2kg potato starch, 0.2kg talcum powder and the 0.1kg Magnesium Stearate of 1kg formula I is obtained tablet according to the ordinary method compacting, comprise the 10mg activeconstituents by every in the tablet of this kind method preparation.

Embodiment F: dragee

According to the method compressed tablets that is similar to embodiment E and carry out dressing according to ordinary method with the Drug coating of sucrose, potato starch, talcum powder, tragakanta and staining agent subsequently.

Embodiment G: capsule

The activeconstituents of 2kg formula I is packed in the hard gelatin capsule according to ordinary method, comprise the 20mg activeconstituents by every capsules of this kind method preparation.

Embodiment H: ampulla

The solution of activeconstituents in the 60L redistilled water of 1kg formula I is carried out sterile filtration, be transferred in the ampoule, freeze-drying and under aseptic condition at no mattress condition lower seal.Each ampoule comprises the 10mg activeconstituents.

Claims (6)

1. be selected from down the compound of group:

And pharmaceutically acceptable derivative, solvate, salt, tautomer and steric isomer, comprise the mixture of their various ratios.

2. contain the described compound of at least a claim 1 and/or its pharmaceutically acceptable derivative, salt, solvate, tautomer and steric isomer, comprise mixture and the optional vehicle and/or the medicine of assistant agent of their various ratios.

3. be selected from down group compound and pharmaceutically acceptable derivative, salt, solvate, tautomer and steric isomer, comprise that the mixture of their various ratios is used for the treatment of and/or resist purposes in the medicine of cancer, tumor growth, metastasis growth, fibrosis, restenosis, HIV infection, alzheimer's disease, atherosclerosis and/or promotion wound healing in preparation:

4. the purposes of claim 3, wherein said tumour is selected from tesselated epithelium knurl, bladder cancer, cancer of the stomach, kidney, head and neck cancer, esophagus cancer, cervical cancer, thyroid carcinoma, intestinal cancer, liver cancer, the cancer of the brain, prostate cancer, genitourinary cancer, lymphsystem cancer, cancer of the stomach, laryngocarcinoma, lung cancer, adenocarcinoma of lung, minicell adenocarcinoma of lung, carcinoma of the pancreas, glioblastoma, colorectal carcinoma, mammary cancer, neoplastic hematologic disorder and immunity system tumour, acute myelocytic leukemia, chronic granulocytic leukemia, kemia, chronic lymphatic leukemia.

5. compound described in the claim 3 and/or the acceptable salt of its physiology and solvate are used for the treatment of purposes in the medicine of solid tumor in preparation, wherein treat the compound described in the claim 3 of significant quantity and are selected from following compound combined administration: 1) estrogenic agents, 2) androgen receptor modifier, 3) retinoid receptor modulators, 4) cytotoxic agent, 5) antiproliferative, 6) prenyl protein transferase inhibitor, 7) HMG CoA-reductase inhibitors, 8) hiv protease inhibitor, 9) reverse transcriptase inhibitors and 10) other angiogenesis inhibitor.

6. compound described in the claim 3 and/or the acceptable salt of its physiology and solvate are used for the treatment of purposes in the medicine of solid tumor in preparation, wherein treat compound and the radiotherapy described in the claim 3 of significant quantity and are selected from following compound combined administration: 1) estrogenic agents, 2) androgen receptor modifier, 3) retinoid receptor modulators, 4) cytotoxic agent, 5) antiproliferative, 6) prenyl protein transferase inhibitor, 7) HMG CoA-reductase inhibitors, 8) hiv protease inhibitor, 9) reverse transcriptase inhibitors and 10) other angiogenesis inhibitor.